In the detergent, personal care and food products industry there is a strong trend towards natural ingredients of these products and to environmentally acceptable production processes. Enzymic conversions are very important for fulfilling these consumer demands, as these processes can be completely natural. Moreover enzymic processes are very specific and consequently will produce minimum amounts of waste products. Such processes can be carried out in water at mild temperatures and atmospheric pressure. However enzymic processes based on free enzymes are either quite expensive due to the loss of enzymes or require expensive equipment, like ultra-membrane systems to entrap the enzyme.
Alternatively enzymes can be immobilized either physically or chemically. The latter method has often the disadvantage that coupling is carried out using non-natural chemicals and in processes that are not attractive from an environmental point of view. Moreover chemical modification of enzymes is nearly always not very specific, which means that coupling can affect the activity of the enzyme negatively. Physical immobilization can comply with consumer demands, however also physical immobilization may affect the activity of the enzyme in a negative way. Moreover, a physically immobilized enzyme is in equilibrium with free enzyme, which means that in continuous reactors, according to the laws of thermodynamics, substantial losses of enzyme are unavoidable.
There are a few publications on immobilization of enzymes to microbial cells (see reference 1). The present invention provides a method for immobilizing enzymes to cell walls of microbial cells in a very precise way. Additionally, the immobilization does not require any chemical or physical coupling step and is very efficient. Some extracellular proteins are known to have special functions which they can perform only if they remain bound to the cell wall of the host cell. Often this type of protein has a long C-terminal part that anchors it in the cell wall. These C-terminal parts have very special amino acid sequences. A typical example is anchoring via C-terminal sequences enriched in proline (see reference 2). Another mechanism to anchor proteins in cell walls is that the protein has a glycosyl-phosphatidyl-inositol (GPI) anchor (see reference 3) and that the C-terminal part of the protein contains a substantial number of potential serine and threonine glycosylation sites. O-Glycosylation of these sites gives a rod-like conformation to the C-terminal part of these proteins. Another feature of these manno-proteins is that they seem to be linked to the glucan in the cell wall of lower eukaryotes, as they cannot be extracted from the cell wall with SDS, but can be liberated by glucanase treatment.